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1.
PLoS One ; 10(4): e0123709, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25856504

RESUMEN

RATIONALE: Recent work in preclinical models suggests that signalling via the pro-angiogenic and pro-inflammatory cytokine, CXCL12 (SDF-1), plays an important pathogenic role in pulmonary hypertension (PH). The objective of this study was to establish whether circulating concentrations of CXCL12α were elevated in patients with PAH and related to mortality. METHODS: Plasma samples were collected from patients with idiopathic pulmonary arterial hypertension (IPAH) and PAH associated with connective tissue diseases (CTD-PAH) attending two pulmonary hypertension referral centres (n = 95) and from age and gender matched healthy controls (n = 44). Patients were subsequently monitored throughout a period of five years. RESULTS: CXCL12α concentrations were elevated in PAH groups compared to controls (P<0.05) and receiver-operating-characteristic analysis showed that plasma CXCL12α concentrations discriminated patients from healthy controls (AUC 0.80, 95% confidence interval 0.73-0.88). Kaplan Meier analysis indicated that elevated plasma CXCL12α concentration was associated with reduced survival (P<0.01). Multivariate Cox proportional hazards model showed that elevated CXCL12α independently predicted (P<0.05) earlier death in PAH with a hazard ratio (95% confidence interval) of 2.25 (1.01-5.00). In the largest subset by WHO functional class (Class 3, 65% of patients) elevated CXCL12α independently predicted (P<0.05) earlier death, hazard ratio 2.27 (1.05-4.89). CONCLUSIONS: Our data show that elevated concentrations of circulating CXCL12α in PAH predicted poorer survival. Furthermore, elevated circulating CXCL12α was an independent risk factor for death that could potentially be included in a prognostic model and guide therapy.


Asunto(s)
Quimiocina CXCL12/sangre , Hipertensión Pulmonar Primaria Familiar/sangre , Pronóstico , Adulto , Anciano , Presión Sanguínea , Hipertensión Pulmonar Primaria Familiar/epidemiología , Hipertensión Pulmonar Primaria Familiar/patología , Femenino , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Modelos de Riesgos Proporcionales , Arteria Pulmonar/metabolismo , Arteria Pulmonar/patología
2.
Am J Respir Crit Care Med ; 186(2): 162-9, 2012 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-22592805

RESUMEN

RATIONALE: Macrophage migration inhibitory factor (MIF) is a proinflammatory mediator with unique tautomerase enzymatic activity; the precise function has not been clearly defined. We previously demonstrated that individual patients with cystic fibrosis (CF) who are genetically predisposed to be high MIF producers develop accelerated end-organ injury. OBJECTIVES: To characterize the effects of the MIF-CATT polymorphism in patients with CF ex vivo. To investigate the role of MIF's tautomerase activity in a murine model of Pseudomonas aeruginosa infection. METHODS: MIF and tumor necrosis factor (TNF)-α protein levels were assessed in plasma or peripheral blood mononuclear cell (PBMC) supernatants by ELISA. A murine pulmonary model of chronic Pseudomonas infection was used in MIF wild-type mice (mif(+/+)) and in tautomerase-null, MIF gene knockin mice (mif (P1G/P1G)). MEASUREMENTS AND MAIN RESULTS: MIF protein was measured in plasma and PBMCs from 5- and 6-CATT patients with CF; LPS-induced TNF-α production from PBMCs was also assessed. The effect of a specific inhibitor of MIF-tautomerase activity, ISO-1, was investigated in PBMCs. In the murine infection model, total weight loss, differential cell counts, bacterial load, and intraacinar airspace/tissue volume were measured. MIF and TNF-α levels were increased in 6-CATT compared with 5-CATT patients with CF. LPS-induced TNF-α production from PBMCs was attenuated in the presence of ISO-1. In a murine model of Pseudomonas infection, significantly less pulmonary inflammation and bacterial load was observed in mif(P1G/P1G) compared with mif(+/+) mice. CONCLUSIONS: MIF-tautomerase activity may provide a novel therapeutic target in patients with chronic inflammatory diseases such as CF, particularly those patients who are genetically predisposed to produce increased levels of this cytokine.


Asunto(s)
Fibrosis Quística/enzimología , Factores Inhibidores de la Migración de Macrófagos/fisiología , Adulto , Alelos , Animales , Fibrosis Quística/sangre , Fibrosis Quística/etiología , Fibrosis Quística/genética , Femenino , Técnicas de Sustitución del Gen , Humanos , Factores Inhibidores de la Migración de Macrófagos/sangre , Masculino , Ratones , Ratones Endogámicos C57BL , Neumonía/sangre , Neumonía/enzimología , Neumonía/etiología , Polimorfismo Genético , Infecciones por Pseudomonas/inmunología , Reacción en Cadena en Tiempo Real de la Polimerasa , Secuencias Repetitivas de Ácidos Nucleicos/genética , Infecciones del Sistema Respiratorio/inmunología , Estudios Retrospectivos , Factor de Necrosis Tumoral alfa/sangre
3.
Circulation ; 125(7): 920-30, 2012 Feb 21.
Artículo en Inglés | MEDLINE | ID: mdl-22247494

RESUMEN

BACKGROUND: Pulmonary hypertension occurs in chronic hypoxic lung diseases, significantly worsening morbidity and mortality. The important role of altered bone morphogenetic protein (BMP) signaling in pulmonary hypertension was first suspected after the identification of heterozygous BMP receptor mutations as the underlying defect in the rare heritable form of pulmonary arterial hypertension. Subsequently, it was demonstrated that BMP signaling was also reduced in common forms of pulmonary hypertension, including hypoxic pulmonary hypertension; however, the mechanism of this reduction has not previously been elucidated. METHODS AND RESULTS: Expression of 2 BMP antagonists, gremlin 1 and gremlin 2, was higher in the lung than in other organs, and gremlin 1 was further increased in the walls of small intrapulmonary vessels of mice during the development of hypoxic pulmonary hypertension. Hypoxia stimulated gremlin secretion from human pulmonary microvascular endothelial cells in vitro, which inhibited endothelial BMP signaling and BMP-stimulated endothelial repair. Haplodeficiency of gremlin 1 augmented BMP signaling in the hypoxic mouse lung and reduced pulmonary vascular resistance by attenuating vascular remodeling. Furthermore, gremlin was increased in the walls of small intrapulmonary vessels in idiopathic pulmonary arterial hypertension and the rare heritable form of pulmonary arterial hypertension in a distribution suggesting endothelial localization. CONCLUSIONS: These findings demonstrate a central role for increased gremlin in hypoxia-induced pulmonary vascular remodeling and the increased pulmonary vascular resistance in hypoxic pulmonary hypertension. High levels of basal gremlin expression in the lung may account for the unique vulnerability of the pulmonary circulation to heterozygous mutations of BMP type 2 receptor in pulmonary arterial hypertension.


Asunto(s)
Hipertensión Pulmonar/etiología , Péptidos y Proteínas de Señalización Intercelular/fisiología , Animales , Proteínas Morfogenéticas Óseas/fisiología , Células Cultivadas , Células Endoteliales/metabolismo , Hipoxia/complicaciones , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intercelular/análisis , Masculino , Ratones , Transducción de Señal , Resistencia Vascular
4.
Eur Respir J ; 39(6): 1415-24, 2012 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-22088972

RESUMEN

Given the critical role that endothelial cell dysfunction plays in the pathogenesis of pulmonary hypertensive diseases, we set out to establish if CXCR7, a receptor for the pro-angiogenic ligand CXCL12, is expressed in the vasculature of human lung diseases and examine its role in mediating CXCL12-induced responses in primary pulmonary human microvascular endothelial cells. Receptor and ligand expression was examined in control and explanted human hypertensive lungs, in human plasma and in hypoxic rodent lungs, by ELISA and immunohistochemical studies. Functional in vitro experiments examined the role of CXCR7 in CXCL12-induced lung microvascular endothelial cell proliferation, migration, and wound regeneration and repair. CXCR7 is elevated in the endothelium of explanted human hypertensive lungs and circulating CXCL12 concentrations are significantly elevated in disease. We demonstrate that alveolar hypoxia similar to that found in lung disease increases CXCR7 expression in the pulmonary endothelium. Furthermore, CXCR7 is the receptor through which endothelial cell regeneration and repair, and proliferation, is mediated, whereas signalling via CXCR4 is essential for chemotactic cell migration. Our findings demonstrate that CXCR7 has a critical but previously unrecognised role to play in endothelial cell proliferation, suggesting that CXCR7-mediated signalling may be functionally important in pulmonary vascular diseases.


Asunto(s)
Quimiocina CXCL12/metabolismo , Hipertensión Pulmonar/metabolismo , Fibrosis Pulmonar Idiopática/metabolismo , Receptores CXCR/metabolismo , Enfermedades Vasculares/metabolismo , Adulto , Animales , Células Cultivadas , Quimiocina CXCL12/sangre , Hipertensión Pulmonar Primaria Familiar , Femenino , Humanos , Hipoxia/metabolismo , Pulmón/química , Pulmón/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Microvasos/metabolismo , Persona de Mediana Edad , Ratas , Ratas Sprague-Dawley , Receptores CXCR/sangre , Receptores CXCR4/metabolismo , Cicatrización de Heridas/fisiología
5.
Respir Res ; 12: 17, 2011 Jan 25.
Artículo en Inglés | MEDLINE | ID: mdl-21266048

RESUMEN

BACKGROUND: Chronic alveolar hypoxia, due to residence at high altitude or chronic obstructive lung diseases, leads to pulmonary hypertension, which may be further complicated by right heart failure, increasing morbidity and mortality. In the non-diseased lung, angiogenesis occurs in chronic hypoxia and may act in a protective, adaptive manner. To date, little is known about the behaviour of individual vascular endothelial growth factor (VEGF) family ligands in hypoxia-induced pulmonary angiogenesis. The aim of this study was to examine the expression of placenta growth factor (PlGF) and VEGFB during the development of hypoxic pulmonary angiogenesis and their functional effects on the pulmonary endothelium. METHODS: Male Sprague Dawley rats were exposed to conditions of normoxia (21% O2) or hypoxia (10% O2) for 1-21 days. Stereological analysis of vascular structure, real-time PCR analysis of vascular endothelial growth factor A (VEGFA), VEGFB, placenta growth factor (PlGF), VEGF receptor 1 (VEGFR1) and VEGFR2, immunohistochemistry and western blots were completed. The effects of VEGF ligands on human pulmonary microvascular endothelial cells were determined using a wound-healing assay. RESULTS: Typical vascular remodelling and angiogenesis were observed in the hypoxic lung. PlGF and VEGFB mRNA expression were significantly increased in the hypoxic lung. Immunohistochemical analysis showed reduced expression of VEGFB protein in hypoxia although PlGF protein was unchanged. The expression of VEGFA mRNA and protein was unchanged. In vitro PlGF at high concentration mimicked the wound-healing actions of VEGFA on pulmonary microvascular endothelial monolayers. Low concentrations of PlGF potentiated the wound-healing actions of VEGFA while higher concentrations of PlGF were without this effect. VEGFB inhibited the wound-healing actions of VEGFA while VEGFB and PlGF together were mutually antagonistic. CONCLUSIONS: VEGFB and PlGF can either inhibit or potentiate the actions of VEGFA, depending on their relative concentrations, which change in the hypoxic lung. Thus their actions in vivo depend on their specific concentrations within the microenvironment of the alveolar wall during the course of adaptation to pulmonary hypoxia.


Asunto(s)
Endotelio Vascular/metabolismo , Hipertensión Pulmonar/metabolismo , Hipoxia/metabolismo , Pulmón/irrigación sanguínea , Pulmón/metabolismo , Neovascularización Fisiológica , Proteínas Gestacionales/metabolismo , Factor B de Crecimiento Endotelial Vascular/metabolismo , Células Epiteliales Alveolares/metabolismo , Animales , Western Blotting , Movimiento Celular , Modelos Animales de Enfermedad , Endotelio Vascular/fisiopatología , Hipertensión Pulmonar/genética , Hipertensión Pulmonar/patología , Hipertensión Pulmonar/fisiopatología , Hipoxia/genética , Hipoxia/patología , Hipoxia/fisiopatología , Inmunohistoquímica , Pulmón/patología , Masculino , Factor de Crecimiento Placentario , Reacción en Cadena de la Polimerasa , Proteínas Gestacionales/genética , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Factores de Tiempo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Factor B de Crecimiento Endotelial Vascular/genética , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo
6.
Am J Respir Cell Mol Biol ; 42(5): 517-23, 2010 May.
Artículo en Inglés | MEDLINE | ID: mdl-19574532

RESUMEN

Gremlin is an extracellular glycoprotein that was first identified over a decade ago through its important role in embryonic development, in which it acts as an antagonist of bone morphogenetic protein actions. It plays a critical role in the development of normal airways and the pulmonary circulation in the embryo. More recently, considerable evidence has been presented for a role for gremlin in the pathogenesis of lung diseases, particularly pulmonary hypertension and idiopathic pulmonary fibrosis. The purpose of this article is to review this evidence, consider the potential mechanisms and multicellular actions by which gremlin contributes to disease pathogenesis, and suggest future avenues of research.


Asunto(s)
Péptidos y Proteínas de Señalización Intercelular/metabolismo , Enfermedades Pulmonares/metabolismo , Pulmón/embriología , Pulmón/patología , Animales , Receptores de Proteínas Morfogenéticas Óseas/metabolismo , Proteínas Morfogenéticas Óseas/antagonistas & inhibidores , Pulmón/metabolismo , Especificidad de Órganos
7.
Am J Respir Crit Care Med ; 178(9): 977-83, 2008 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-18689465

RESUMEN

RATIONALE: Pulmonary hypertension is a common complication of chronic hypoxic lung diseases and is associated with increased morbidity and reduced survival. The pulmonary vascular changes in response to hypoxia, both structural and functional, are unique to this circulation. OBJECTIVES: To identify transcription factor pathways uniquely activated in the lung in response to hypoxia. METHODS: After exposure to environmental hypoxia (10% O(2)) for varying periods (3 h to 2 wk), lungs and systemic organs were isolated from groups of adult male mice. Bioinformatic examination of genes the expression of which changed in the hypoxic lung (assessed using microarray analysis) identified potential lung-selective transcription factors controlling these changes in gene expression. In separate further experiments, lung-selective activation of these candidate transcription factors was tested in hypoxic mice and by comparing hypoxic responses of primary human pulmonary and cardiac microvascular endothelial cells in vitro. MEASUREMENTS AND MAIN RESULTS: Bioinformatic analysis identified cAMP response element binding (CREB) family members as candidate lung-selective hypoxia-responsive transcription factors. Further in vivo experiments demonstrated activation of CREB and activating transcription factor (ATF)1 and up-regulation of CREB family-responsive genes in the hypoxic lung, but not in other organs. Hypoxia-dependent CREB activation and CREB-responsive gene expression was observed in human primary lung, but not cardiac microvascular endothelial cells. CONCLUSIONS: These findings suggest that activation of CREB and AFT1 plays a key role in the lung-specific responses to hypoxia, and that lung microvascular endothelial cells are important, proximal effector cells in the specific responses of the pulmonary circulation to hypoxia.


Asunto(s)
Proteína de Unión a Elemento de Respuesta al AMP Cíclico/genética , Hipertensión Pulmonar/fisiopatología , Hipoxia/fisiopatología , Pulmón/fisiopatología , Factores de Transcripción/genética , Factor de Transcripción Activador 1/genética , Animales , Hipoxia de la Célula , Células Cultivadas , Modelos Animales de Enfermedad , Células Endoteliales , Regulación de la Expresión Génica/genética , Humanos , Hipertensión Pulmonar/genética , Hipoxia/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Factores de Tiempo , Resistencia Vascular/genética
8.
Am J Physiol Lung Cell Mol Physiol ; 295(2): L272-84, 2008 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-18469115

RESUMEN

Pulmonary hypoxia is a common complication of chronic lung diseases leading to the development of pulmonary hypertension. The underlying sustained increase in vascular resistance in hypoxia is a response unique to the lung. Thus we hypothesized that there are genes for which expression is altered selectively in the lung in response to alveolar hypoxia. Using a novel subtractive array strategy, we compared gene responses to hypoxia in primary human pulmonary microvascular endothelial cells (HMVEC-L) with those in cardiac microvascular endothelium and identified 90 genes (forming 9 clusters) differentially regulated in the lung endothelium. From one cluster, we confirmed that the bone morphogenetic protein (BMP) antagonist, gremlin 1, was upregulated in the hypoxic murine lung in vivo but was unchanged in five systemic organs. We also demonstrated that gremlin protein was significantly increased by hypoxia in vivo and inhibited HMVEC-L responses to BMP stimulation in vitro. Furthermore, significant upregulation of gremlin was measured in lungs of patients with pulmonary hypertensive disease. From a second cluster, we showed that CXC receptor 7, a receptor for the proangiogenic chemokine CXCL12, was selectively upregulated in the hypoxic lung in vivo, confirming that our subtractive strategy had successfully identified a second lung-selective hypoxia-responsive gene. We conclude that hypoxia, typical of that encountered in pulmonary disease, causes lung-specific alterations in gene expression. This gives new insights into the mechanisms of pulmonary hypertension and vascular loss in chronic lung disease and identifies gremlin 1 as a potentially important mediator of vascular changes in hypoxic pulmonary hypertension.


Asunto(s)
Proteínas Morfogenéticas Óseas/antagonistas & inhibidores , Hipertensión Pulmonar/metabolismo , Hipoxia/metabolismo , Péptidos y Proteínas de Señalización Intercelular/biosíntesis , Alveolos Pulmonares/metabolismo , Mucosa Respiratoria/metabolismo , Animales , Proteínas Morfogenéticas Óseas/metabolismo , Hipoxia de la Célula , Células Cultivadas , Quimiocina CXCL12/biosíntesis , Modelos Animales de Enfermedad , Células Endoteliales/metabolismo , Regulación de la Expresión Génica , Humanos , Ratones , Alveolos Pulmonares/irrigación sanguínea , Receptores CXCR/biosíntesis , Receptores Acoplados a Proteínas G/biosíntesis , Mucosa Respiratoria/irrigación sanguínea
9.
PLoS Med ; 2(8): e199, 2005 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-16107186

RESUMEN

BACKGROUND: The differential pathophysiologic mechanisms that trigger and maintain the two forms of inflammatory bowel disease (IBD), Crohn disease (CD), and ulcerative colitis (UC) are only partially understood. cDNA microarrays can be used to decipher gene regulation events at a genome-wide level and to identify novel unknown genes that might be involved in perpetuating inflammatory disease progression. METHODS AND FINDINGS: High-density cDNA microarrays representing 33,792 UniGene clusters were prepared. Biopsies were taken from the sigmoid colon of normal controls (n = 11), CD patients (n = 10) and UC patients (n = 10). 33P-radiolabeled cDNA from purified poly(A)+ RNA extracted from biopsies (unpooled) was hybridized to the arrays. We identified 500 and 272 transcripts differentially regulated in CD and UC, respectively. Interesting hits were independently verified by real-time PCR in a second sample of 100 individuals, and immunohistochemistry was used for exemplary localization. The main findings point to novel molecules important in abnormal immune regulation and the highly disturbed cell biology of colonic epithelial cells in IBD pathogenesis, e.g., CYLD (cylindromatosis, turban tumor syndrome) and CDH11 (cadherin 11, type 2). By the nature of the array setup, many of the genes identified were to our knowledge previously uncharacterized, and prediction of the putative function of a subsection of these genes indicate that some could be involved in early events in disease pathophysiology. CONCLUSION: A comprehensive set of candidate genes not previously associated with IBD was revealed, which underlines the polygenic and complex nature of the disease. It points out substantial differences in pathophysiology between CD and UC. The multiple unknown genes identified may stimulate new research in the fields of barrier mechanisms and cell signalling in the context of IBD, and ultimately new therapeutic approaches.


Asunto(s)
Enfermedades Inflamatorias del Intestino/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Antígenos CD/genética , Antígenos CD/metabolismo , Moléculas de Adhesión Celular/genética , Moléculas de Adhesión Celular/metabolismo , Colitis Ulcerosa/genética , Colitis Ulcerosa/metabolismo , Enfermedad de Crohn/genética , Enfermedad de Crohn/metabolismo , Enzima Desubiquitinante CYLD , Femenino , Perfilación de la Expresión Génica/métodos , Regulación de la Expresión Génica , Genoma Humano , Humanos , Inmunohistoquímica , Enfermedades Inflamatorias del Intestino/metabolismo , Mucosa Intestinal/metabolismo , Masculino , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteína Quinasa C/genética , Proteína Quinasa C/metabolismo , Proteína Quinasa C beta , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo
10.
Circ Res ; 97(2): 185-91, 2005 Jul 22.
Artículo en Inglés | MEDLINE | ID: mdl-15961717

RESUMEN

Pulmonary hypertension (PH) is a common complication of chronic hypoxic lung diseases, which increase morbidity and mortality. Hypoxic PH has previously been attributed to structural changes in the pulmonary vasculature including narrowing of the vascular lumen and loss of vessels, which produce a fixed increase in resistance. Using quantitative stereology, we now show that chronic hypoxia caused PH and remodeling of the blood vessel walls in rats but that this remodeling did not lead to structural narrowing of the vascular lumen. Sustained inhibition of the RhoA/Rho-kinase pathway throughout the period of hypoxic exposure attenuated PH and prevented remodeling in intra-acinar vessels without enlarging the structurally determined lumen diameter. In chronically hypoxic lungs, acute Rho kinase inhibition markedly decreased PVR but did not alter the alveolar to arterial oxygen gap. In addition to increased vascular resistance, chronic hypoxia induced Rho kinase-dependent capillary angiogenesis. Thus, hypoxic PH was not caused by fixed structural changes in the vasculature but by sustained vasoconstriction, which was largely Rho kinase dependent. Importantly, this vasoconstriction had no role in ventilation-perfusion matching and optimization of gas exchange. Rho kinase also mediated hypoxia-induced capillary angiogenesis, a previously unrecognized but potentially important adaptive response.


Asunto(s)
Amidas/farmacología , Inhibidores Enzimáticos/farmacología , Hipertensión Pulmonar/etiología , Hipoxia/fisiopatología , Pulmón/irrigación sanguínea , Neovascularización Fisiológica/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Piridinas/farmacología , Animales , Presión Sanguínea/efectos de los fármacos , Capilares/efectos de los fármacos , Capilares/fisiopatología , Hipertensión Pulmonar/prevención & control , Hipoxia/complicaciones , Péptidos y Proteínas de Señalización Intracelular , Masculino , Consumo de Oxígeno/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/fisiología , Ratas , Factor A de Crecimiento Endotelial Vascular/genética , Resistencia Vascular/efectos de los fármacos , Vasoconstricción/efectos de los fármacos , Quinasas Asociadas a rho , Proteína de Unión al GTP rhoA/análisis
11.
BMC Bioinformatics ; 6: 37, 2005 Feb 23.
Artículo en Inglés | MEDLINE | ID: mdl-15727680

RESUMEN

BACKGROUND: Normalization is the process of removing non-biological sources of variation between array experiments. Recent investigations of data in gene expression databases for varying organisms and tissues have shown that the majority of expressed genes exhibit a power-law distribution with an exponent close to -1 (i.e. obey Zipf's law). Based on the observation that our single channel and two channel microarray data sets also followed a power-law distribution, we were motivated to develop a normalization method based on this law, and examine how it compares with existing published techniques. A computationally simple and intuitively appealing technique based on this observation is presented. RESULTS: Using pairwise comparisons using MA plots (log ratio vs. log intensity), we compared this novel method to previously published normalization techniques, namely global normalization to the mean, the quantile method, and a variation on the loess normalization method designed specifically for boutique microarrays. Results indicated that, for single channel microarrays, the quantile method was superior with regard to eliminating intensity-dependent effects (banana curves), but Zipf's law normalization does minimize this effect by rotating the data distribution such that the maximal number of data points lie on the zero of the log ratio axis. For two channel boutique microarrays, the Zipf's law normalizations performed as well as, or better than existing techniques. CONCLUSION: Zipf's law normalization is a useful tool where the Quantile method cannot be applied, as is the case with microarrays containing functionally specific gene sets (boutique arrays).


Asunto(s)
Biología Computacional/métodos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Algoritmos , Análisis de Varianza , Animales , Calibración , Simulación por Computador , Interpretación Estadística de Datos , Bases de Datos Genéticas , Etiquetas de Secuencia Expresada , Expresión Génica , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Funciones de Verosimilitud , Modelos Genéticos , Modelos Estadísticos , Modelos Teóricos , ARN/química , Ratas , Análisis de Regresión , Sensibilidad y Especificidad , Análisis de Secuencia de ADN
12.
Nat Genet ; 36(5): 476-80, 2004 May.
Artículo en Inglés | MEDLINE | ID: mdl-15107852

RESUMEN

Crohn disease and ulcerative colitis are two subphenotypes of inflammatory bowel disease (IBD), a complex disorder resulting from gene-environment interaction. We refined our previously defined linkage region for IBD on chromosome 10q23 and used positional cloning to identify genetic variants in DLG5 associated with IBD. DLG5 encodes a scaffolding protein involved in the maintenance of epithelial integrity. We identified two distinct haplotypes with a replicable distortion in transmission (P = 0.000023 and P = 0.004 for association with IBD, P = 0.00012 and P = 0.04 for association with Crohn disease). One of the risk-associated DLG5 haplotypes is distinguished from the common haplotype by a nonsynonymous single-nucleotide polymorphism 113G-->A, resulting in the amino acid substitution R30Q in the DUF622 domain of DLG5. This mutation probably impedes scaffolding of DLG5. We stratified the study sample according to the presence of risk-associated CARD15 variants to study potential gene-gene interaction. We found a significant difference in association of the 113A DLG5 variant with Crohn disease in affected individuals carrying the risk-associated CARD15 alleles versus those carrying non-risk-associated CARD15 alleles. This is suggestive of a complex pattern of gene-gene interaction between DLG5 and CARD15, reflecting the complex nature of polygenic diseases. Further functional studies will evaluate the biological significance of DLG5 variants.


Asunto(s)
Cromosomas Humanos Par 10/genética , Colitis Ulcerosa/genética , Enfermedad de Crohn/genética , Variación Genética , Péptidos y Proteínas de Señalización Intracelular , Proteínas de la Membrana/genética , Proteínas Supresoras de Tumor/genética , Alelos , Sustitución de Aminoácidos , Proteínas Portadoras/genética , Estudios de Casos y Controles , Niño , Mapeo Cromosómico , Clonación Molecular , Estudios de Cohortes , Colitis Ulcerosa/epidemiología , Enfermedad de Crohn/epidemiología , Europa (Continente)/epidemiología , Femenino , Genotipo , Haplotipos , Humanos , Masculino , Proteína Adaptadora de Señalización NOD2 , Linaje , Polimorfismo de Nucleótido Simple , Factores de Riesgo
13.
Physiol Genomics ; 16(3): 361-70, 2004 Feb 13.
Artículo en Inglés | MEDLINE | ID: mdl-14645736

RESUMEN

Large-scale public data mining will become more common as public release of microarray data sets becomes a corequisite for publication. Therefore, there is an urgent need to clarify whether data from different microarray platforms are comparable. To assess the compatibility of microarray data, results were compared from the two main types of high-throughput microarray expression technologies, namely, an oligonucleotide-based and a cDNA-based platform, using RNA obtained from complex tissue (human colonic mucosa) of five individuals. From 715 sequence-verified genes represented on both platforms, 64% of the genes matched in "present" or "absent" calls made by both platforms. Calls were influenced by spurious signals caused by Alu repeats in cDNA clones, clone annotation errors, or matched probes that were designed to different regions of the gene; however, these factors could not completely account for the level of call discordance observed. Expression levels in sequence-verified, platform-overlapping genes were not related, as demonstrated by weakly positive rank order correlation. This study demonstrates that there is only moderate overlap in the results from the two array systems. This fact should be carefully considered when performing large-scale analyses on data originating from different microarray platforms.


Asunto(s)
ADN Complementario/genética , Análisis de Secuencia por Matrices de Oligonucleótidos/instrumentación , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Oligonucleótidos/genética , Anciano , Elementos Alu/genética , Biopsia , Colon/metabolismo , Femenino , Perfilación de la Expresión Génica/instrumentación , Perfilación de la Expresión Génica/métodos , Humanos , Mucosa Intestinal/metabolismo , Masculino , Persona de Mediana Edad , ARN Mensajero/análisis , ARN Mensajero/genética , Reproducibilidad de los Resultados
14.
Eur J Surg Suppl ; (587): 70-6, 2002.
Artículo en Inglés | MEDLINE | ID: mdl-16144204

RESUMEN

Genomic techniques offer new approaches to the investigation of the aetiology and pathophysiology of intestinal disorders. An important field that is relevant to treatment is the pharmacogenetic investigation of gene variations that may predict response to certain drugs in order to target these drugs more precisely. For the surgeon the perioperative situation will be here of great interest. To date only about 8000 of the estimated 30,000-50,000 human genes have been characterised, so the use of techniques for global analysis of gene expression may allow the identification of new pathways or molecules. In can be anticipated that genomic methods will profoundly influence the treatment of gastrointestinal disorders and will lead to new insights into both aetiology and pathophysiology of chronic intestinal inflammation.


Asunto(s)
Enfermedades Gastrointestinales/tratamiento farmacológico , Enfermedades Gastrointestinales/fisiopatología , Farmacogenética , Biblioteca de Genes , Variación Genética , Humanos , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/fisiopatología , Análisis de Secuencia por Matrices de Oligonucleótidos , Factor de Transcripción STAT6/metabolismo
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